Simulated Body Fluid Research Articles

Bioactive material‑sodium alginate-polyvinyl alcohol composite film scaffold for bone tissue engineering application

Road accidents and infection-causing diseases during bone surgery are serious problems in orthopedics, and thus, addressing these pressing challenges is crucial. In the present study, the 70S30C calcium silicate bioactive material (BM) is synthesized by a sustainable approach employing a precipitation method using recycled rice husk and eggshells as a precursor of silica and calcium. Further, 70S30C BM is composited with sodium alginate (SA) and polyvinyl alcohol (PVA), and the films were prepared by solvent casting method. The composite films were prepared without the addition of acid, binder, and crosslinking agents. Further, the films were characterized by BET, XRD, ATR-FTIR, SEM, and EDS mapping. The in vitro bioactivity and biodegradation study is performed in the simulated body fluid (SBF). The in vitro haemolysis study is executed using human blood and the results demonstrate haemocompatibility of the composite films. The ex ovo CAM assay also exhibits good neovascularization. The in vitro and in vivo biocompatibility assay proves its non-toxic nature. Further, the in vivo study reveals that the engineered composite film demonstrates accelerated osteogenesis. This work broadens the orthopedic potential of the composite film and offers bioactivity, haemocompatibility, angiogenesis, non-toxicity, and in vivo osteogenesis which would serve as a potential candidate for bone tissue engineering application.

Duplex Fluorinated and Atomic Layer Deposition-Derived ZrO2 Coatings Improve the Corrosion Resistance and Mechanical Properties of Mg-2Zn-0.46Y-0.5Nd (wt.%) Alloy Plates and Screws.

This study investigated the corrosion resistance and mechanical properties of Mg-2Zn-0.46Y-0.5Nd (wt.%) alloy plates and screws with fluorinated coatings and atomic layer deposition (ALD)-derived zirconia (ZrO2) coatings in vitro under physiological stress conditions. Synthetic polyurethane hemimandible replicas were split and fixed as the following three groups of magnesium alloy plates and screws: no additional surface coating treatment (Group A), with fluorinated coatings (Group B), and with duplex fluorinated and ALD-derived 100 nm ZrO2 coatings (Group C). A circulating stress of 1-10 N was applied to the distal bone segment, and a 4-week simulated body fluid immersion test was employed to study the remaining material volume and the mechanical properties of the different groups. Compared with Group A and Group B, the degradation rate of magnesium alloy plates and screws' head regions was significantly slowed down under the protection of duplex MgF2/ZrO2 coatings (p < 0.01). There was no significant difference in the degradation rate of the screw shaft region between groups (p = 0.077). In contrast to fluoride coatings, duplex MgF2/ZrO2 coatings maintained the mechanical strength of magnesium alloy plates and screws after a 14 day in vitro SBF immersion test. We conclude that duplex MgF2/ZrO2 coatings exhibited a certain protective effect on the Mg alloy plates and screws under physiological stress conditions.

Maleic Anhydride-Derived Copolymers Conjugated with Beta-Lactam Antibiotics: Synthesis, Characterization, In Vitro Activity/Stability Tests with Antibacterial Studies

The synthesis and characterization of biocompatible three different maleic anhydride co-polymer conjugated with two different beta-lactam antibiotics at in vitro conditions were conducted. The polymer–drug conjugates were synthesized by coupling β-lactam antibiotics via amide bonds to the copolymer. In this work, six different drug-functionalized maleic anhydride copolymers (DFMACs) were synthesized by the chemical conjugation method. This method is based on the ring-opening reaction of the anhydride ring of the copolymer to form an amide bond linking the drug. The synthesized DFMACs were characterized by 1H NMR and FTIR/ATR spectroscopies and analyses were carried out by UV/VIS spectroscopy and Zeta-sizer instrument in detail with consecutive antibacterial tests. The existence of a newly formed amide covalent bond between the drug and the copolymer chains was confirmed by 1H NMR and FTIR/ATR studies. This is the first report on the application of the selected branched biodegradable polymeric matrices for the covalent conjugation of ampicillin and cefalexin. Optimum stability and activity conditions for the synthesized DFMACs were determined. Analyses were conducted under in vitro conditions including varying pH values and simulated body fluids as a function of time to obtain new drug delivery system candidates for the two different antibiotics.

Cellulose Acetate and Polycaprolactone Fibre Coatings on Medical-Grade Metal Substrates for Controlled Drug Release.

This study explores a method that has the potential to be cost effective in inhibiting biofilm formation on metallic prostheses, thereby preventing rejection or the requirement for replacement. A cost-effective metal alloy used in biomedical implants was chosen as the substrate, and ibuprofen (Ibu), a well-known anti-inflammatory drug, was selected for drug release tests for its widespread availability and accessibility. Multilayer coatings consisting of cellulose acetate (CA), polycaprolactone (PCL), and chitosan (CHI), with or without ibuprofen (Ibu) content, were applied onto medical-grade stainless steel (SS-316 type) through electrospinning, electrospray, or blow spinning. The adhesion of the CA, PCL, and layered CA/PCL membranes, with thicknesses ranging from 20 to 100 μm, to SS substrates varied between 0.15 N and 0.22 N without CHI, which increased to 0.21 and 0.74 N, respectively, when a CHI interlayer was introduced by electrospraying between the SS and the coatings. Although drug release in a simulated body fluid (SBF) medium is predominantly governed by diffusion-driven mechanisms in all single- and multilayer coatings, a delayed release was noted in CA coatings containing Ibu when overlaid with a PCL coating produced by blow spinning. This suggests avenues for further investigations into combinations of multilayer coatings, both with and without drug-imbued layers.

Optimized in vitro assessment of ZrO2–CaO/PMMA hybrid biocomposite with multi-walled carbon nanotube reinforcement for enhanced bone reconstruction

In this study, multi-walled carbon nanotubes (MWCNTs) are integrated into a poly (methyl methacrylate) (PMMA) and zirconium oxide (ZrO2) biocomposite, stabilized with calcium oxide (CaO). This research aims to pave the way for further optimization of the biocomposite for targeted applications in bone tissue engineering. The incorporation of MWCNTs is intended to enhance the mechanical properties and bioactivity of the composite, making it a suitable candidate for bone reconstruction. Comprehensive analyses were conducted using field emission scanning electron microscopy (FESEM) and energy dispersive X-ray spectroscopy (EDS) to characterize the structural and chemical changes in the biocomposite during immersion in simulated body fluid (SBF). These analyses revealed a significant formation of a robust apatite layer on the composite surface after three days of immersion. Notably, the rate of apatite formation accelerated with the incorporation of MWCNTs, indicating an enhancement in the bioactivity of the composite. The study findings demonstrate that an MWCNT-reinforced PMMA/ZrO2–CaO composite exhibits excellent biocompatibility, as well as accelerated bioactivity. These properties are crucial for bone tissue engineering applications, where materials must integrate seamlessly with natural bone and support new bone formation. The results confirm the potential of this advanced biocomposite as a promising biomaterial for bone replacement procedures, offering improved performance over traditional materials.

Derivatization, solid-phase microextraction and gas chromatography for determination of dialdehydes—biomarkers of oxidative stress

Dialdehydes are often used as biomarkers, molecules of interest for diagnosis, medical studies, and scientific research. A well-known example is malondialdehyde, which is a biomarker of oxidative stress formed during lipid peroxidation. We used combination of derivatization with O-(1,2,3,4,5)-pentafluorobenzylhydroxylamine, headspace solid-phase microextraction and gas chromatography with flame ionization detector for the analysis of dialdehydes. This combination allows to ensure the stability of analytes by derivatization, to separate them from the matrix, to preconcentrate by solid-phase microextraction, and to perform determination of dialdehydes. All the parameters affecting sensitivity and selectivity (pH and time of derivatization, sorption time, temperature of sorption, salt addition) were studied specifically for malondialdehyde, glyoxal, and methylglyoxal. The addition of salting-out agent resulted in decrease in the signal; therefore, a possible matrix effect was tested using simulated body fluid as model solution with ion composition similar to blood plasma. Established conditions for this method for dialdehydes showed 84–102% recovery in the model solution and allowed to reach the limits of quantification 0.2–0.8 μmol/L, which is low enough for the analysis of real samples. This method has the potential to be automated and is environmentally friendly because it does not require organic solvents and single-used filters or cartridges.

Electrochemical properties and corrosion induced mechanical degradation of biomedical Zn–Cu–Li alloy in simulated body fluid

The electrochemical behavior and corrosion induced mechanical degradation of biodegradable Zn–2Cu–0.03Li alloy were in detail investigated. The results indicated that the corrosion product film presented an increasing corrosion resistance, while the localized corrosion induced by microgalvanic effect was intensified. Corrosion pits, as the typical defect, acted as the crack initiation sites and accelerated the fracture process of the Zn–2Cu–0.03Li alloy. Accordingly, the fracture mode transformed from ductile to brittle failure. Significant loss in ultimate tensile strength and elongation of the alloys occurred due to strong stress concentration at corrosion pit, threatening the mechanical reliability after implantation.

Structure and Properties of Bioactive Titanium Dioxide Surface Layers Produced on NiTi Shape Memory Alloy in Low-Temperature Plasma.

The NiTi alloy, known for its shape memory and superelasticity, is increasingly used in medicine. However, its high nickel content requires enhanced biocompatibility for long-term implants. Low-temperature plasma treatments under glow-discharge conditions can improve surface properties without compromising mechanical integrity. This study explores the surface modification of a NiTi alloy by oxidizing it in low-temperature plasma. We examine the impact of process temperatures and sample preparation (mechanical grinding and polishing) on the structure of the produced titanium oxide layers. Surface properties, including topography, morphology, chemical composition, and bioactivity, were analyzed using TEM, SEM, EDS, and an optical profilometer. Bioactivity was assessed through the deposition of calcium phosphate in simulated body fluid (SBF). The low-temperature plasma oxidization produced titanium dioxide layers (29-55 nm thick) with a predominantly nanocrystalline rutile structure. Layer thickness increased with extended processing time and higher temperatures (up to 390 °C), though the relationship was not linear. Higher temperatures led to thicker layers with more precipitates and inhomogeneities. The oxidized layers showed increased bioactivity after 14 and 30 days in SBF. Low-temperature plasma oxidation produces bioactive titanium oxide layers on NiTi alloys, with a structure and properties that can be tuned through process parameters. This method could enhance the biocompatibility of NiTi alloys for medical implants.

Synthesis of Heterostructured TiO2 Nanopores/Nanotubes by Anodizing at High Voltages.

This paper reports on the coating of heterostructured TiO2 nanopores/nanotubes on Ti substrates by anodizing at high voltages to design surfaces for biomedical implants. As the anodized voltage from 50 V to 350 V was applied, the microstructure of the coating shifted from regular TiO2 nanotubes to heterostructured TiO2 nanopores/nanotubes. In addition, the dimension of the heterostructured TiO2 nanopores/nanotubes was a function of voltage. The electrochemical characteristics of TiO2 nanotubes and heterostructured TiO2 nanopores/nanotubes were evaluated in simulated body fluid (SBF) solution. The creation of heterostructured TiO2 nanopores/nanotubes on Ti substrates resulted in a significant increase in BHK cell attachment compared to that of the Ti substrates and the TiO2 nanotubes.

Investigation of the structural properties of Si4+-doped HAP coatings on Ti-6Al-4V substrate as a corrosion barrier in biomedical media

The coatings of co-substituted bioactives play an important role in stimulating biological and physical properties for biomedical applications. Hydroxyapatite (HAP), a mineral compound possessing a hexagonal structure, exhibits high thermodynamic stability under typical physiological conditions. In this experimental study, Si4+-doped HAP coatings with varying concentrations (2.5 %, 5 %, and 10 % wt) were prepared on a Ti-6Al-4V substrate using the spin-coating technique and extreme centrifugal force (2500 rpm). The coated samples were characterized by XRD, FTIR, AFM, FESEM, and EDS techniques. The presence of cracks and voids in the HAP coatings detrimentally affected the performance of the implants. Tetraethyl orthosilicate (TEOS) was used in the coatings to mitigate the cracks and voids. Polarization and EIS studies in the simulated body fluid (SBF) were used to assess the corrosion resistance of the coatings. The decrease in current density and the improved resistance to corrosion observed in the coated samples, as compared to the uncoated Ti-6Al-4V, demonstrate the corrosion-inhibiting effects of anodizing. The integration of Si4+ ions in the structure of HAP caused the size of nanoparticles to decrease from 16.23 nm to 11.67 nm, and the surface roughness of the produced coatings exhibited a decrease from 29.18 to 5.65 nm. In the Nyquist plots, the 5 % Si4+-doped HAP coating displayed the highest Rct value of 17.99 MΩ.cm2 compared to the bare Ti-6Al-4V with a resistance of 0.13 MΩ.cm2. The 5 % Si4+-doped HAP coating was chosen as the optimal sample based on the study results. The results of this work show that surface modification with Si4+-doped HAP coatings is a promising approach for improving the functionality of the Ti-6Al-4 V alloy for biomedical applications.

Multiscale investigation on the formation path of the apatite phase in bioactive glasses

Mesoporous bioactive glasses (MBGs) have been reported as promising biomaterials to stimulate and promote the growth of bones. When exposed to Simulated Body Fluid (SBF), MBGs develop an apatite phase called carbonate hydroxyapatite (H(C)A) on their surfaces that closely mimics the mineral phase of bones. In order to gain deeper insights into the key stages of the surface reactions of two different types of MBGs (58S and 70S30C) soaked in SBF, we have used high energy total X-ray scattering coupled to Pair Distribution Function (PDF) analysis, along with Raman spectroscopy and scanning electron microscopy. This type of coupled analytical approach can ultimately help to unravel the details of the reaction kinetics in the complex calcium phosphate environment around the BGs surfaces. The data obtained in our in-vitro study point to the formation of other intermediate phases like amorphous calcium phosphate (ACP) and calcite (CC), that transform after a specific time into H(C)A depending on the composition of the MBGs, their surface properties and their interaction times with SBF.

Degradation Behavior of Medical MgZZC-1 in Various Simulated Body Fluids.

Magnesium-based biodegradable metal bone implants exhibit superior mechanical properties compared to biodegradable polymers for orthopedic and cardiovascular stents. In this study, MgZZC-x (x = 1, 1.2) alloys were screened by in vitro biocompatibility tests in three simulated body fluids under nontoxic conditions. The MgZZC-1 alloys with better biocompatibility were selected to predict the days required for complete degradation. The evolution of degradation products was analyzed, and the mechanism of formation of the product film was inferred. A degradation kinetic model was established to investigate the effect of MEM components on the degradation of the alloys. The results demonstrate that the proteins in MEM can greatly retard the degradation progress by attaching to the surface of MgZZC-1 alloys, which are predicted to degrade completely within 341 days. The carbonate and phosphate buffers were adjusted to pH in MEM solution, delaying the degradation of magnesium alloys. This process in MEM more accurately reflects the actual degradation in the body and is superior to that in Hanks and SBF solutions. This study will promote the application of biodegradable materials in clinical medicine.

Investigations on the corrosion characteristics of additive manufactured Mg–Ag alloy in simulated body fluids for biodegradable implants

Investigations on the corrosion characteristics of additive manufactured Mg–Ag alloy in simulated body fluids for biodegradable implants

Efficient Bioactive Surface Coatings with Calcium Minerals: Step-Wise Biomimetic Transformation of Vaterite to Carbonated Apatite.

Carbonated apatite (CAp), known as the main mineral that makes up human bone, can be utilized in conjunction with scaffolds to increase their bioactivity. Various methods (e.g., co-precipitation, hydrothermal, and biomimetic coatings) have been used to provide bioactivity by forming CAp on surfaces similar to bone minerals. Among them, the use of simulated body fluids (SBF) is the most popular biomimetic method for generating CAp, as it can provide a mimetic environment. However, coating methods using SBF require at least a week for CAp formation. The long time it takes to coat biomimetic scaffolds is a point of improvement in a field that requires rapid regeneration. Here, we report a step-wise biomimetic coating method to form CAp using calcium carbonate vaterite (CCV) as a precursor. We can manufacture CCV-transformed CAp (V-CAp) on the surface in 4 h at least by immersing CCV in a phosphate solution. The V-CAp deposited surface was analyzed using scanning electron microscopy (SEM) images according to the type of phosphate solutions to optimize the reaction conditions. X-ray diffraction (XRD) and attenuated total reflection-Fourier transform infrared (ATR-FTIR) analysis validated the conversion of CCV to V-CAp on surfaces. In addition, the bioactivity of V-CAp coating was analyzed by the proliferation and differentiation of osteoblasts in vitro. V-CAp showed 2.3-folded higher cell proliferation and 1.4-fold higher ALP activity than the glass surface. The step-wise method of CCV-transformed CAp is a biocompatible method that allows the environment of bone regeneration and has the potential to confer bioactivity to biomaterial surfaces, such as imparting bioactivity to non-bioactive metal or scaffold surfaces within one day. It can rapidly form carbonated apatite, which can greatly improve time efficiency in research and industrial applications.

Enhancing bioactivity, radiopacity, and structural properties of borophosphate glass through barium oxide modification and controlled thermal treatment

This study explores the impact of replacing P2O5 and B2O3 with varying amounts of barium oxide (BaO, 0–10 mol%) in borophosphate glasses. The investigation focuses on the structural, bioactivity, and radiopaque properties of the resulting materials. Four glass samples were prepared using a melt-quench technique. Their bioactivity was assessed through immersion in simulated body fluid (SBF) for 28 days. Additionally, a 24-h thermal treatment at 530 °C was employed to induce crystallization and evaluate the bioactivity of the resulting glass-ceramics. Initial X-ray diffraction (XRD) analysis confirmed the amorphous nature of the glasses. Conversely, crystalline phases were identified after the thermal treatment. Increasing BaO content led to higher volumetric density while altering the molar volume and structure as revealed by Fourier-transform infrared spectroscopy (FTIR) analysis. Lower pH values were observed with increasing BaO content, suggesting its influence on controlling the release of ions from the material. Both XRD and FTIR analyses detected the formation of hydroxyapatite (HAp) and fluorapatite (FAp) phases in all samples. Notably, the glass-ceramic containing 10 mol% BaO exhibited the most significant apatite formation. Radiopacity measurements improved with increasing BaO content, indicating potential applications in bone replacement materials due to enhanced visibility in X-ray imaging. Cell viability tests confirmed the non-toxic nature of the samples, with higher BaO content promoting cell proliferation, suggesting potential for tissue regeneration. Overall, these findings suggest promising biomedical applications for all the studied compositions.

Enhancing wear resistance, anti-corrosion property and surface bioactivity of a beta-titanium alloy by adopting surface mechanical attrition treatment followed by phosphorus ion implantation

A two-step method of surface mechanical attrition treatment (SMAT) followed by phosphorous (P) ion-implantation was adopted to treat Ti-25Nb-3Mo-2Sn-3Zr (named as TLM) alloy. The initial SMAT process refined the grains from micrometer-scale to about 10–30 nm, and begot the TLM surface much rougher with apparently improved hydrophilicity. The subsequent P ion-implantation led to the formation of a hybrid structure of the TiP, β-Ti nanograins and P-interstitial amorphous in the surface layer of the TLM alloy, and resulted in a slight decrease of the surface roughness and wettability of the SMAT-processed alloy. The indentation, tribology and potentiodynamic polarization tests indicated that the SMAT procedure concurrently enhanced the microhardness, wear-resisting property and corrosion resistance of the TLM alloy, and the followed P-ion implantation could further improve these tested properties. Moreover, the P-ion implanted TLM alloy was verified to exhibit much better surface bioactivity (including biomineralization in the simulated body fluid (SBF) and in-vitro cell adhesion) as compared to the original TLM alloy. With the remarkable improvements of the wear resistance and anti-corrosion property as well as the surface bioactivity of the P ion-implanted TLM alloy, our study provides an alternative avenue to fabricating new-type Ti-based orthopedic implants with superior comprehensive performance.

Fabrication and characterization of hydroxyapatite coatings on anodized magnesium alloys by electrochemical and chemical methods intended for biodegradable implants

The fabrication of hydroxyapatite (HAP)/MgO composite coatings on Mg alloy is crucial for enhancing the corrosion resistance and biocompatibility of biomedical implants. In this study, we aimed to investigate the effects of two different surface modification methods, i.e., electrochemical (electrodeposition, ED) and chemical (solution treatment, ST), on the phase structure, degradation properties, and biocompatibility of the composite coatings in comparison to the anodized coating. The surface morphologies and crystalline structures of the coatings were characterized by scanning electron microscopy (SEM) and X-ray diffraction (XRD). Subsequently, the degradation rate of the coatings in simulated body fluid were comprehensively evaluated by using electrochemical impedance spectroscopy (EIS), potentiodynamic polarization and scanning electrochemical microscopy (SECM) tests. Additionally, in-vitro cell proliferation assays were employed to quantitatively assess the biocompatibilities of the coatings. The results showed that both ED and ST methods were effective in depositing HAP on anodized Mg alloy, resulting in different surface morphologies with hydroxyapatite layer thicknesses of 2.71 μm and 3.56 μm, respectively. In this way, the HAP-deposited coatings exhibited improved corrosion resistances and biocompatibilities compared with those of the anodized coating. Specifically, the ST-deposited composite film displayed superior degradability and biocompatibility which was attributed to its filiform surface morphology and thicker HAP layer. Overall, the present study demonstrates the potential of HAP/MgO composite coatings for biomedical applications, with implications for the development of advanced implant materials with improved performance and biocompatibility.

Electrochemical Evaluation of Biomedical Titanium and Its Alloys in Simulated Body Fluids with Inflammation Condition H2O2

Electrochemical Evaluation of Biomedical Titanium and Its Alloys in Simulated Body Fluids with Inflammation Condition H2O2

Porous silica-doped calcium phosphate scaffolds prepared via in-situ foaming method

The effect of silica (SiO2) addition (0 wt%-20 wt%) on the microstructural and mechanical properties, as well as the in vitro response of calcium phosphate scaffolds for potential application in bone tissue engineering (BTE) was investigated in this research. Scaffolds characterized by high porosity (77%–88 %) and interconnected spherical pores with a broad range of pore sizes (5–600 μm) were fabricated using in-situ foaming method. Incorporated silica affected the phase transformation of hydroxyapatite (HA) to β-tricalcium phosphate (β-TCP) and led to the development of new crystalline silica-rich phases like silicocarnotite and wollastonite. The reinforcement of silica became apparent during the tests of mechanical properties. Scaffolds with 5 wt% of SiO2 exhibited compressive strength (1.13 MPa) higher than pure HA scaffolds (0.93 MPa). Bone bonding potential of the materials was tested in simulated body fluid (SBF), demonstrating this potential in silica-doped samples. Additionally, degradation experiments showed gradual material degradation, making it suitable for BTE applications. Furthermore, cell culture studies using human mesenchymal stromal cells (MSC) confirmed the scaffold's non-toxicity and provided insights into how the silica content influences cell viability, morphology, and osteogenic potential. The findings of this study offer valuable insights into the design and development of advanced scaffolds with tailored properties for effective BTE applications.

Dual-layered carbonated hydroxyapatite/polypyrrole: A novel strategy for bone fracture repair implants

This study presents the fabrication of a carbonate-doped hydroxyapatite/polypyrrole thin film on 304 stainless steel (CHA/PPY/304 SS) using a cyclic voltammetry electrochemical method to enhance bone fracture repair. The in-situ substitution mechanism was evaluated through estimated reaction energy, and the impact of surface modification on structural properties was examined using the Rietveld refinement method. Following the electrodeposition of CHA on PPY-coated 304 SS, a distinctive flower-like structure emerged, with average cluster and pore sizes of 3 ± 1 μm and 130 ± 10 nm, respectively. Rietveld refinement analysis revealed A-type carbonate substitution, indicated by an increase in the a parameter and a reduction in the c parameter compared to the standard. Subsequent deposition of the bilayer coating on 304 SS significantly improved adhesion strength to 10.7 ± 0.2 MPa, along with increased hydrophilicity and Vickers hardness. Notably, enhanced bioactivity was observed, evidenced by thickened nanoflake-like structures and the formation of island-like apatite deposits after immersion in simulated body fluid (SBF) for 7 days. This bilayer coating shows promise for surface modification of metallic implants and tissue engineering scaffolds.